[Extracellular toxins of Pseudomonas aeruginosa. I. Purification and characterization of two exoproteases (author's transl)].
Alkaline protease (protease I) and a protease with elastase activity (protease II) were isolated from two different strains of Pseudomonas aeruginosa (PA). Proteolytic activity was measured during the early exponential phase of growth and was highest when cultures reached the stationary growth phase. The extracellular character of protease I and II was demonstrated by measuring the intra- and extracellular ATP-concentration. Purification was achieved by precipitation with 65% ammonium sulfate, precipitation with 70% acetone, gelfiltration on Sephadex G-100 and chromatography on DEAE-Sephacel. The purified proteases were characterized. The pH for optimal proteolytic activity of protease I was at pH 9--10, for protease II at pH 8--9. Both enzymes cleaved casein and gelatine, in addition protein II elastin. Enzymatic activity of protease II was inhibited by 10(-3) M EDTA at pH 8.1 to 82%. Inactivation of protease I was not achieved by 10(-2) M EDTA. Molecular weight of protease I was estimated at 57,000, molecular weight of protease II at 39,000. Both enzymes consist of one polypeptide chain. In isoelectric focusing the protease I was separated into two components with pH values of 8.5 and 8.7, while protease II had isoelectric points of pH 6.0 and 6.4. Further characterization of protease I was done with amino acid analysis. Protease I was fairly stable over a pH range of 6--9 at room temperature. The optimal temperature for proteolytic activity was 60 degrees C. The results are discussed in view of proteases of other PA-strains.
D000596:Amino Acids / Q000032:analysis; D004492:Edetic Acid / Q000494:pharmacology; D020689:Exopeptidases; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D008970:Molecular Weight; D010196:Pancreatic Elastase / Q000378:metabolism; D010447:Peptide Hydrolases / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000254:growth & development; D013696:Temperature
D000596:Amino Acids; D004492:Edetic Acid; D020689:Exopeptidases; D010447:Peptide Hydrolases; D010196:Pancreatic Elastase
Extrazelluläre Toxine von Pseudomonas aeruginosa. I. Reinigung und Charakterisierung zweier Exoproteasen.